Compositions and methods for correcting limb girdle muscular dystrophy type 2C using exon skipping

Abstract

The invention is directed to one or more antisense polynucleotides and their use in pharmaceutical compositions in a strategy to induce exon skipping in the γ-sarcoglycan gene in patients suffering from Limb-Girdle Muscular Dystrophy-2C (LGM-D2C) or in patients at risk of such a disease. The invention also provides methods of preventing or treating muscular dystrophy, e.g., LGMD2C, by exon skipping in the gamma sarcoglycan gene using antisense polynucleotides. Accordingly, in some aspects the invention provides an isolated antisense oligonucleotide, wherein the oligonucleotide specifically hybridizes to an exon target region of a γ-sarcoglycan RNA. In another aspect, the invention provides a method of inducing exon-skipping of a gamma sarcoglycan RNA, comprising delivering an antisense oligonucleotide or a composition to a cell.

What is claimed is: 
     
       1. A modified antisense oligonucleotide (AON) selected from the group consisting of oligonucleotides having a sequence as set out in SEQ ID NOs: 4-9, 11-12, 14-29, and 31-34. 
     
     
       2. The antisense oligonucleotide of  claim 1 , wherein the oligonucleotide cannot form an RNase H substrate. 
     
     
       3. The antisense oligonucleotide of  claim 1 , comprising a modified oligonucleotide backbone. 
     
     
       4. The antisense oligonucleotide of  claim 3 , wherein the modified oligonucleotide backbone comprises a modified moiety substituted for the sugar. 
     
     
       5. The antisense oligonucleotide of  claim 4 , wherein the modified moiety is a Morpholino. 
     
     
       6. The antisense oligonucleotide of  claim 3 , wherein the modified oligonucleotide backbone comprises at least one modified internucleotide linkage. 
     
     
       7. The antisense oligonucleotide of  claim 6 , wherein the modified internucleotide linkage is a tricyclo-DNA (tc-DNA) modification. 
     
     
       8. The antisense oligonucleotide of  claim 6 , wherein the modified internucleotide linkage comprises a modified phosphate. 
     
     
       9. The antisense oligonucleotide of  claim 8 , wherein the modified phosphate is selected from the group consisting of a methyl phosphonate, a methyl phosphorothioate, a phosphoromorpholidate, a phosphoropiperazidate and a phosphoroamidate. 
     
     
       10. The antisense oligonucleotide of  claim 3 , wherein the oligonucleotide is a 2′-O-methyl-oligoribonucleotide. 
     
     
       11. The antisense oligonucleotide of  claim 1 , wherein the oligonucleotide comprises a peptide nucleic acid. 
     
     
       12. The antisense oligonucleotide of  claim 1 , wherein the oligonucleotide is chemically linked to one or more conjugates that enhance the activity, cellular distribution, or cellular uptake of the antisense oligonucleotide. 
     
     
       13. The antisense oligonucleotide of  claim 12 , wherein the oligonucleotide is chemically linked to a polyethylene glycol molecule. 
     
     
       14. The antisense oligonucleotide of  claim 12  wherein the conjugate is a peptide that enhances cellular uptake. 
     
     
       15. The antisense oligonucleotide of  claim 14  wherein the peptide is selected from the group consisting of a nuclear localization signal (NLS), HIV-1 TAT protein, a peptide comprising an integrin binding domain, oligolysine, adenovirus fiber protein and a peptide comprising a receptor-mediated endocytosis (RME) domain. 
     
     
       16. A method of inducing exon-skipping of a gamma sarcoglycan RNA, comprising delivering to a cell the antisense oligonucleotide of  claim 1 , thereby inducing exon-skipping of the gamma sarcoglycan RNA. 
     
     
       17. The method of  claim 16 , wherein the cell is a human muscle cell. 
     
     
       18. A kit comprising the modified antisense oligonucleotide of  claim 1 , optionally in a container, and a package insert, package label, instructions or other labeling. 
     
     
       19. The kit of  claim 18 , further comprising an additional modified oligonucleotide, wherein the additional modified oligonucleotide specifically hybridizes to an exon in a gamma sarcoglycan RNA. 
     
     
       20. The composition of  claim 1 , further comprising a physiologically compatible buffer. 
     
     
       21. The method of  claim 16 , wherein the cell is a human muscle cell. 
     
     
       22. The method of  claim 21 , wherein the human muscle cell is in a patient. 
     
     
       23. The method of  claim 22 , wherein the patient has muscular dystrophy. 
     
     
       24. The method of  claim 23 , wherein the muscular dystrophy is Limb Girdle Muscular Dystrophy type 2C (LGMD2C).